THE ROLE OF CSF-1 MEDIATED MACROPHAGE CHEMOTAXIS IN CARCINOMA CELL INVASION

CSF-1介导的巨噬细胞趋化作用在癌细胞侵袭中的作用

• 批准号: 7035701
• 负责人: Dianne Cox
• 金额: $ 30.91万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7035701
• 关键词: cell cell interaction; cell cycle; cell differentiation; gene therapy; macrophage; protein structure function

DESCRIPTION (provided by applicant): Tumor-associated macrophages (TAM), which are present in large numbers in many tumors, appear to play an important role in promoting the progression of solid tumors to an invasive, metastatic phenotype. It has been shown recently that TAM participate in a paracrine loop with carcinoma cells such that the CSF-1 - producing carcinoma cells and the EGF-secreting macrophages (MF) interact to promote mutual chemotaxis leading to invasion and extravasation of carcinoma cells. In MFs, PI 3-kinase, Cdc42 and WASP (Wiskott- Aldrich syndrome protein) are required for migrating cells to detect the source of a chemoattractant. It has been speculated that amplification of the extracellular gradient occurs through an intracellular positive feedback loop involving PI 3-kinase, Rho GTPases and actin assembly. The precise function of WASP in gradient detection is not known. Based on preliminary data, WASP activity is required for CSF-1 induced actin polymerization in MFs which may contribute to the reinforcement of the positive feedback loop in CSF- 1 gradient detection (chemotactic sensing) leading to efficient chemotaxis. In the first Specific Aim, the role of PI3K and Cdc42 in the activation of WASP will be determined using PI3K inhibitors, CSF-1 R mutations that lack PI3K activation, and by reducing endogenous levels of Cdc42 using siRNA technology. The mechanisms of WASP activation will be examined through mutational analysis of a fluorescence resonance energy transfer (FRET) based WASP biosensor. In Specific Aim 2, the role of WASP mediated actin polymerization in chemotactic sensing will be determined. The role of WASP in the localization of PI3K and Cdc42 in chemotactic sensing will be determined by live cell imaging. In addition, we will perform biochemical isolation of CSF-1 elicited pseudopods from wild-type and WASP-deficient macrophages in order to identify potential WASP interacting proteins. In Specific Aim 3. The effect of WASP on polarization and movement of MF and carcinoma cells will be examined using an in vitro assay that reconstitutes the paracrine interaction between MF and tumor cells. Relevance: Understanding how MFs migrate into a tumor site and their interaction with carcinoma cells, is an important area of investigation in cancer biology. Since WASP is specifically required for MF chemotaxis it may represent a novel target and may lead to new therapies to prevent metastasis

描述（由申请人提供）：肿瘤相关巨噬细胞（TAM）在许多肿瘤中大量存在，似乎在促进实体瘤进展为侵袭性、转移性表型方面发挥着重要作用。最近的研究表明，TAM 参与癌细胞的旁分泌环路，使得产生 CSF-1 的癌细胞和分泌 EGF 的巨噬细胞 (MF) 相互作用，促进相互趋化性，导致癌细胞的侵袭和外渗。在 MF 中，迁移细胞需要 PI 3-激酶、Cdc42 和 WASP（Wiskott-Aldrich 综合征蛋白）来检测趋化剂的来源。据推测，细胞外梯度的放大是通过涉及 PI 3-激酶、Rho GTPases 和肌动蛋白组装的细胞内正反馈环发生的。 WASP 在梯度检测中的精确功能尚不清楚。根据初步数据，WASP 活性是 CSF-1 诱导 MF 中肌动蛋白聚合所必需的，这可能有助于加强 CSF-1 梯度检测（趋化传感）中的正反馈回路，从而实现有效的趋化性。在第一个具体目标中，将使用 PI3K 抑制剂、缺乏 PI3K 激活的 CSF-1 R 突变以及使用 siRNA 技术降低 Cdc42 的内源水平来确定 PI3K 和 Cdc42 在 WASP 激活中的作用。将通过基于 WASP 生物传感器的荧光共振能量转移 (FRET) 的突变分析来检查 WASP 激活机制。在具体目标 2 中，将确定 WASP 介导的肌动蛋白聚合在趋化传感中的作用。 WASP 在趋化传感中 PI3K 和 Cdc42 定位中的作用将通过活细胞成像确定。此外，我们将从野生型和 WASP 缺陷型巨噬细胞中对 CSF-1 引发的伪足进行生化分离，以鉴定潜在的 WASP 相互作用蛋白。在具体目标 3 中，将使用体外测定来检查 WASP 对 MF 和癌细胞的极化和运动的影响，该测定重建 MF 和肿瘤细胞之间的旁分泌相互作用。相关性：了解 MF 如何迁移到肿瘤部位及其与癌细胞的相互作用，是癌症生物学研究的一个重要领域。由于 WASP 是 MF 趋化性所必需的，因此它可能代表一个新的靶标，并可能导致预防转移的新疗法